I’ve heard several reports now that labs can sometimes see some kind of either Oct4-GFP reporter activity or pluripotency gene expression in acid treated cells, but the scientists do not seem particularly encouraged.

One said that the cells gave a glimmer of interesting gene expression, but would not grow at all and s/he thought the gene expression and Oct4-GFP may have been manifestations of cell death leading to weird chromatin decondensation events.

Another lab had cells growing after acid treatment but only saw at best the faintest, transient sign of pluripotency indicators. The cells did not look “normal” whatever that means.

So overall either the cells with even slightly encouraging signs wouldn’t grow or the cells that would grow were not encouraging in terms of seeming at all like pluripotent stem cells.

By far the most common message I’m getting from people, however, is that they’ve simply given up trying to replicate STAP stem cells at least until the detailed methods paper on STAP stem cells comes out. I think some have given up entirely.

If we give STAP stem cells a year to be reproduced as Dr. Wakayama has suggested and which seems reasonable to me, I would argue that it is still important that labs working on the method communicate, share data if they are comfortable doing so, and keep a critical eye on all of this.

Question:
1. Are Pasteur pipettes glass or plastic?http://www.thomassci.com/scientific-supplies/Pasteur-Pipets
2. The name of the company of Pasteur pipettes and type of pipettes (size, form) or manually produced?
3. As for mechanical trituration, she mentioned as follows. Surprisingly, she showed
5~20% appearance of STAP cells by this unsteady procedures repeating trituration for 20 minutes!. She did not describe the time of trituration in low PH treatment.
For “rigorous trituration”, what speed of trituration?

“To give a mechanical stress to mature cells, a pasture pipette was heated and then stretched to create thin capillaries with the lumens approximately 50 mm in diameter, and then broken into appropriate lengths. Mature somatic cells were then repeatedly triturated through these pipettes for 20 min, and then cultured for 7 days.”

“Discussion
A remaining question is whether cellular reprogramming is initiated specifically by the low-pH treatment or also by some other types of sublethal stress such as physical damage, plasma membrane perforation, osmotic pressure shock, growth-factor deprivation, heat shock or high Ca2+ exposure. At least some of these stressors, particularly physical damage by rigorous trituration and membrane perforation by streptolysin O, induced the generation of Oct4-GFP1 cells from CD451 cells (Extended Data Fig. 9a; see Methods).”

It was recently found out that there seem to be two further critical points of discussion. Since both of them are mainly circulated in Japanese only right now, I’ve tried to summarize them below in English. Note that I am a researcher outside this field, and I am not a native speaker of English or Japanese, so I might not be using correct terms. As a researcher, I am simply deeply concerned with all the recent fuss on the sloppiness of supporting data on sensational claims.

It was pointed that teratoma formation including a seemingly complete pancreas (with even a lobule) is rather infeasible. Some also points that they can observe lacteals, which again is considered infeasible for teratoma formation.

2., Sequencing data

A person who analyzed the published sequencing data (http://slashdot.jp/journal/578529) reported that STAP cells and CD45+s treated under the low-pH environment exhibit no trace of TCR rearrangement. It seems to contradict with an common observation that TCR rearrangement should happen if STAP cells are indeed made by reprogramming T cells, as claimed by Obokata et al. This analysis on the sequencing data is in fact confirmed in the recently published protocol by Riken (the person who did this analysis claims that s/he submitted his analysis to Riken before this protocol is published):

“(iii) We have established multiple STAP stem cell lines from STAP cells derived from CD45+ haematopoietic cells. Of eight clones examined, none contained the rearranged TCR allele, suggesting the possibility of negative cell-type-dependent bias (including maturation of the cell of origin) for STAP cells to give rise to STAP stem cells in the conversion process. This may be relevant to the fact that STAP cell conversion was less efficient when non-neonatal cells were used as somatic cells of origin in the current protocol.”http://www.cdb.riken.jp/jp/04_news/articles/pdf/14/protocol_exchange_v1.pdf

I by no means try to (nor I can) make any conclusion. I leave any further analysis to experts here, as they seem worth some attentions from experts all over the world.

Last, but not least, let me iterate and emphasize again that your effort on supporting crowd sourcing academic verifications is really respectful, regardless of how this incident turns out (thanks for your sincere response to my comment back then, too).

I think we are living in an fantastic world right now as researchers – lots of people have accesses to latest research work, actively discuss them, and freely publish their opinions and analyses, and all of these can happen even within a month after publication!